Interpretive Summary: Chronic alcohol consumption causes bad quality of bone and easier bone fracture, the mechanisms of how chronic alcohol effect on bone are largely unknown and potential treatments to prevent alcohol-induced bone fracture remain unclear. We have previously demonstrated that, chronic ethanol (EtOH) treatment leads to accumulation of reactive oxygen species (ROS) in bone cells dependent on one enzyme called NADPH (nicotinamide adenine dinucleotide phosphate) oxidase (Nox). EtOH-induced ROS production might effect on both inhibition of bone formation and enhancing of bone resorption. Using a cycling female Sprague-Dawley rat model, we gave alcohol directly into gastrointestinal track. We found that EtOH infusion for 4 weeks reduced bone quality. Co-administration of diphenylene iodonium chloride (DPI), a Nox inhibitor, by daily injection of 1 mg/kg/d, abolished EtOH-induced inhibition of bone formation. EtOH-induced inhibition of bone formation was associated with up-regulation of mRNA levels of all three Nox subtypes 1, 2, 4 in bone. These data suggest that inhibition of Nox expression or activity may be new target for prevention or treatment of chronic EtOH-induced bone loss, and perhaps other conditions resulting in oxidative stress associated bone resorption such as aging.